Velo cardio facial syndrome and DiGeorge syndrome (VCFS/DGS) are the most common human developmental disorders that result from haploinsufficiency. Most of the patients are hemizygous for 3 Mb deletion on human chromosome 22q11. Patients with VCFS/DGS have a number of abnormalities in tissues and organ systems that are derived from the neural crest. These include facial dysmorphology, cardiovascular defects, immune system defects and many others. Using the mouse as a model system the PIs have been able to show that mice that either overexpress or have reduced expression of Tbx1, a gene located in the deleted interval, have defects in many of the organs systems affected in VCFS/DGS patients. TBX1 encodes a transcription factor that is critical for normal neural crest development. Although information about the mechanisms of action of TBX1 is emerging, no systematic efforts to identify the pathways and networks in which TBX1 acts have been undertaken. The PIs propose to do so. TBX1 is highly conserved during evolution and mutations of this gene in the zebrafish, designated van gogh (vgo), develop phenotypes that are similar to those seen in VCFS/DGS patients. Based on these results, the PIs now propose a comparative genetics analysis of the TBX1 pathways and networks in zebrafish and mice. They propose to use gene expression profiling of normal and mutant zebrafish and mouse embryos at the appropriate developmental stages to discover genes whose expression patterns are altered as a result of reduction or absence of TBX1 protein. They will use whole mount in situ hybridization methods to examine the spatiotemporal patterns of expression of a large number of these genes in zebrafish. A subset will also be examined in Tbx1 mutant zebrafish and mice. Genes whose expression patterns match that of TBX1 or in appropriate cells and tissues will be subjected to genetic analysis in zebrafish through overexpression and morpholino-based knock-downs. If mutants for some of these genes are available, they will be examined alone and in combination with TBX1 mutants. In some cases the PIs will generate null and conditional mutant alleles in mice to examine their role in development. The combination of the use of zebrafish and mouse together with high throughput genetic and genomic approaches promise to provide rich information about the pathways and networks in which TBX1 operates.